title: Cloning, expression and characterization of glycoside hydrolase family 11 endoxylanase from Bacillus pumilus ARA.
abstract: An endoxylanase gene, xynA, was cloned from Bacillus pumilus ARA and expressed in Escherichia coli. The open reading frame of the xynA gene was 687 bp encoding a signal peptide and a mature xylanase with a molecular mass of 23 kDa. The enzyme was categorized as a glycosyl hydrolase family 11 member based on the sequence analysis of the putative catalytic domain. The recombinant XynA (Bpu XynA) was purified to homogeneity by Ni-NTA and ion exchange chromatography on DEAE-Sepharose FF. The enzyme exhibited highest activity at pH 6.6 and 50°C. The purified Bpu XynA was stable for at least 2 h at 45°C, and retained over 50% residual activity after being incubated at 60°C for 1 h. The activity of the xylanase was not significantly affected by metal ions and EDTA. The K ( m ) and K ( cat ) /K ( m ) of Bpu XynA for oat-spelt xylan were 5.53 mg/ml and 10.14 ml/mg s at 50°C and pH 6.6. The main product of hydrolysis by Bpu XynA was xylooligosaccharide. The results revealed that the consumption of grass xylan by B. pumilus ARA depended on the synergistic reactions of Bpu XynA and Bpu arabinosidase, and that a typical GH11 xylanase e.g. Tla XynA had capability to remove the side chain of xylan. The properties Bpu XynA make it promising for application in the production of Bifidobacterium growth-promoting factors and in feed industry.
authors.affiliation: Research Center for Biotechnology and Biomass Energy, Nanjing Normal University, Nanjing, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Qu
authors.forename: Weilan
authors.initials: W
authors.lastname: Shao
mesh_terms: D020121:5' Untranslated Regions; D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D020536:Enzyme Activators / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames; D021382:Protein Sorting Signals; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D013997:Time Factors; D014990:Xylans / Q000378:metabolism
chemical_list: D020121:5' Untranslated Regions; D004269:DNA, Bacterial; D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D014990:Xylans; D043364:Endo-1,4-beta Xylanases